Microorganisms possessing a cell-surface component known as an endotoxic lipopolysaccharide (LPS) have been identified as the etiologic agents in a wide variety of diseases. Of particular interest are strains of the genera Neisseria, and Chlamydia, which are associated with certain venereal diseases; the incidence of these diseases having reached almost epidemic proportions
The LPS molecule has been the subject of intense study. See for example, Westphal et al , Bacterial Lipopolysaccharides, Methods of Carbohydrate Chemistry, Vol. V, Academic Press, (1965), pgs 83-91; and Galanos et. al., Biochemistry of Lipids II, T. W. Goodwin (Ed), Univ. Park Press, (1977), Vol. 14, at page 239.
The structure of the lipopolysaccharide has been described in studies of Gram-negative bacteria such as, Escherichia coli and Salmonella typhimurium. In general, the antigen can be visualized as possessing three component regions. Proximal to and imbedded into the outer portion of the cell membrane is the so-called lipid A component. This component has been associated with the endotoxin properties of the molecule and is believed to be a highly conserved sequence; that is to say, a wide variety of organisms would likely possess identical lipid A regions. The middle region of the LPS molecule is another conserved region, the so-called core oligosaccharide. The core oligosaccharide is bipartite in structure based upon the types of sugar moieties of which it is composed. The innermost region which is adjacent to the lipid A component contains the unusual sugar, 3-deoxy-manno-octulosonic acid, also known as ketodeoxyoctanoic acid (KDO). The outermost region of the core structure is comprised of a region of seven-carbon sugars (heptoses) which is followed distally by a region containing six-carbon sugars (hexoses). The most distal region of the LPS molecule is the so-called somatic or (0) region comprised of highly-variable polysaccharide components.
It is this outermost region which induces the strongest immune response in infected organisms and it is to this region that antisera produced thereby is directed. Thus, even though a number of different taxa may possess the same Lipid A and core components of the LPS molecules, because of the antigenic variability of the somatic region, antisera produced thereto will be specific for each type of inducing organism.
Microorganisms can thus be classified into chemotypes based upon their carbohydrate composition in the 0 region of the LPS. One method is described by Merrick in U.S. Pat. No. 3,891,508. Such a composition analysis although somewhat useful for purposes of identification is limited by not being able to discriminate between two organisms which may possess the same kinds of sugars but in different proportions or in different sequences.
The LPS molecule when used as an immunogen, may produce certain toxic side-effects due to the biological activity of the molecule. Several patents, such as U.S. Pat. Nos. 4,185,090 and 4,057,685 relate to methods of reducing the toxicity of the molecule but retaining the antigenicity of the O region. The specificity of the O region in permitting the identification, of only a single species is disclosed by Wallace et. al. in U.S. Pat. No. 4,115,543 relating to the identification of Neisseria gonorrhoeae. Other approaches attempt to find other antigenic inducers and eliminate the LPS molecule from the immunizing mixture as for example, described by Ayme in U.S. Pat. No. 4,337,243.
It has been recently demonstrated that conventional polyclonal sera raised to Salmonella typhimuruin Re mutants cross-react with various Chlamydia species (Nurminen, et al., Science 220: 1279-81 (1983)). There is no suggestion of the diagnostic utility nor of the wider reactive potential of the antibodies as produced and described hereinbelow.
Human monoclonals of the IgG.sub.1 subclass have been demonstrated to be reactive with a limited class of microorganisms namely members of the Chlamydia genera. Although, not particularly well-characterized, the antigen to which the monoclonals are directed is referred to as a lipoprotein complex. Thus, it is probably closely related to the Chlamydia group specific protein as characterized by Caldwell, et al., (J. Immuno. 115: 963-68 (1975)) and clearly distinct from the endotoxic-glycolipid as described herein.
In an interesting, but unrelated application, (PCT application No. WO80/001109) the LPS molecule itself has been used as a marker reagent; being chemically attached to a ligand, it permits the detection of the LPS-ligand complex by standard LPS assay method (The Limulus amebocyte assay).
None of the above references relate to the object of the subject invention, that being, the generation of immunological reagents useful in the identification of a diverse group of LPS producing microorganisms.